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1. ABSTRACT
Edible vaccines offer cost-effective, easily administrable, storable and widely
acceptable as bio friendly particularly in developing countries. Oral administration of
edible vaccines proves to be promising agents for reducing the incidence of various
diseases like hepatitis and diarrhea especially in the developing world, which face the
problem of storing and administering vaccines. Edible vaccines are obtained by
incorporating a particular gene of interest into the plant, which produces the desirable
encoded protein. Edible vaccines are specific to provide mucosal activity along with
systemic immunity. Various foods that are used as alternative agents for injectable
vaccines include cereals (wheat, rice, corn) fruits (bananas) and vegetables (lettuce,
potatoes, tomatoes). Thus, edible vaccines overcome all the problems associated with
traditional vaccines and prove to be best substitutes to traditional vaccines.
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2. INTRODUCTION
A vaccine is a biological preparation that provides active acquired immunity to a
particular disease. A vaccine typically contains an agent that resembles a disease-
causing microorganism and is often made from weakened or killed forms of the
microbe, its toxins, or one of its surface proteins. The agent stimulates the
body's immune system to recognize the agent as a threat, destroy it, and to further
recognize and destroy any of the microorganisms associated with that agent that it
may encounter in the future. Vaccines can be prophylactic (example: to prevent or
ameliorate the effects of a future infection by a natural or "wild" pathogen),
or therapeutic (e.g., vaccines against cancer are being investigated)(Melief, van
Hall et al. 2015).
The administration of vaccines is called vaccination. Vaccination is the most
effective method of preventing infectious diseases; widespread immunity due to
vaccination is largely responsible for the worldwide eradication of smallpox and
the restriction of diseases such as polio, measles, and tetanus from much of the
world. The effectiveness of vaccination has been widely studied and verified; for
example, the influenza vaccine (Chen, Cheng et al. 2016), the HPV vaccine
(Situma 2013), and the chicken poxvaccine (Liesegang 2009). The World Health
Organization (WHO) reports that licensed vaccines are currently available for
twenty-five different preventable infections (Organization 2013).
The terms vaccine and vaccination are derived from Variolae vaccinae (smallpox
of the cow), the term devised by Edward Jenner to denote cowpox. He used it in
1798 in the long title of his Inquiry into the Variolae vaccinae known as the Cow
Pox, in which he described the protective effect of cowpox against smallpox.
2.1. Types of Vaccines:
There are severalbasic types of vaccines. Some vaccines are describedhere:
1. Attenuated whole-agent vaccines use living but attenuated (weakened)
microbes. Examples of attenuated vaccines are the Sabin polio vaccine and those
used against measles, mumps and rubella (MMR). The widely used vaccine
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against the tuberculosis bacillus and certain of the newly introduced, orally
administered typhoid vaccines contain attenuated bacteria.
2. Inactivated whole-agent vaccines use microbes that have been killed.
Inactivated virus vaccines used in humans include those against rabies, influenza
and polio (the Salk polio vaccine). Inactivated bacterial vaccines include those for
pneumococcal pneumonia, cholera, pertussis (whooping cough) and typhoid.
3. Toxoids which are inactivated toxins, are vaccines directed at the toxins
produced by a pathogen. Examples: Vaccines against tetanus and diphtheria.
4. Subunit vaccines use only those antigenic fragments of a microorganism that
best stimulate an immune response. Subunit vaccines that are produced by genetic
modification techniques, meaning that other microbes are programmed to produce
the desired antigenic fraction, are called recombinant vaccines. For example, the
vaccine against the hepatitis В virus consists of a portion of the viral protein coat
that is produced by genetically modified yeast.
5. Conjugated vaccines have been developed in recent years to deal with the poor
immune response of children. The polysaccharides are combined with proteins
such as diphtheria toxoid. This approach has led to the very successful vaccine for
hemophilic influenza type b, which gives significant protection.
6. Nucleic acid vaccines or DNA vaccines are among the newest and most
promising vaccines, although they have not yet resulted in any commercial
vaccine for humans. Experiments with animals show that plasmids of “naked”
DNA injected into muscle results in the production of the protein encoded in the
DNA.
These proteins stimulate an immune response. A problem with this type of
vaccine is that the DNA remains effective only until it is degraded. Indications are
that RNA, which could replicate in the recipient, might be a more effective agent.
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3. CONCEPTS OF EDIBLE VACCINES
Development of edible vaccines involves the process of incorporating the selected
desired genes into plants and then enabling these altered plants to produce the
encoded proteins. This process is known as transformation, and the altered plants
are known as transgenic plants. Edible vaccines like traditional subunit vaccines
consist of antigenic proteins and are devoid of pathogenic genes. Despite this
advantage, traditional subunit vaccines are unaffordable and technology-intensive,
require purification, refrigeration and produce poor mucosal response. Unlikely,
edible vaccines would eliminate the need for trained medical personnel required
for oral administration particularly in children. Production of edible vaccines is
effective process and can be easily scaled up. Edible vaccines offer numerous
advantages like they possess good genetic and heat stability and do not need cold-
chain maintenance. Edible vaccines can be stored at the site of use thus avoiding
long-distance transportation. Syringes and needles are also not required, thus
reduces the incidence of various infections (Webster, Thomas et al. 2002).
Important advantage of edible vaccines is elimination of contamination with
animal viruses-like the mad cow disease, which is a hazard in vaccines developed
from cultured mammalian cells, as plant viruses cannot infect humans. Edible
vaccines act by stimulating the mucosal as well as systemic immunity, as soon
they meet the digestive tract lining. This dual mechanism of action of edible
vaccines provide first-line defence against pathogens attacking via mucosa,
like Mycobacterium tuberculosis and carriers causing diarrhoea, pneumonia,
STDs, HIV etc (Poland, Murray et al. 2002). Oral administration of edible
vaccines to mothers might prove to be useful in immunizing the fetus-in-utero by
transplacental movement of maternal antibodies or the infant through breast-
feeding. Edible vaccines enable the process of seroconversion in the presence of
maternal antibodies, thus playing a possible role in protecting children against
diseases like group-B Streptococcus, respiratory syncytial virus (RSV), etc. At
present edible vaccines are produced for various human and animal diseases
(measles, cholera, foot and mouth disease and hepatitis B, C and E). They can also
be used to prevent exceptional diseases like dengue, hookworm, rabies, etc. by
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combining with other vaccination programmes enabling multiple antigen delivery.
Various foods under investigation for use in edible vaccines include banana,
potato, tomato, lettuce, rice, etc.(Giddings, Allison et al. 2000)
3.1. Developing an Edible Vaccine
The selected gene obtained from the microbes encoding specific antigen can be
handled in two different ways:
1. Suitable plant virus is genetically engineered to produce the desired
peptides/proteins. The recombinant virus is then incorporated into the plant,
which enables it to produce a huge number of new plants from which chimeric
virions are isolated and purified. The consequential edible plant vaccine can
then be used for immunological applications.
2. In another method, the desirable gene is incorporated with plant vector by
transformation. Many other approaches have been utilized which can be
categorized into following groups:
3.1.1. Agrobacterium mediated gene transfer
In this method, the suitable gene (recombinant DNA) is incorporated into the T-
region of a disarmed Ti plasmid of Agrobacterium; a plant pathogen, which is co-
cultured with the plant cells, or tissues that needs to be transformed (Figure 1). This
approach is slow with lower yield however; it showed satisfactory results in
dicotelydenous plants like potato, tomato and tobacco. Researches in some fields
have proven this approach good in expressing the desirable traits by selected genes
in several experimental animals and plants (Mercenier, Wiedermann et al. 2001;
Chikwamba, Cunnick et al. 2002).
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Figure 1: Development of edible vaccines from potato.
3.1.2. Biolistic method
This sophisticated method involves the use of gene gun that fires the gene containing
DNA coated metal (e.g. gold, tungsten) particles at the plant cells (Taylor and
Fauquet 2002). Plant cells are then permitted to grow in new plants, which are later
on cloned to produce ample number of crop with similar genetic composition. This
approach is highly attractive due to its undependability on regeneration ability of the
species as DNA is directly incorporated into cells of plant. However, requirement of
expensive device particle gun adds to the major drawback to this method.
3.1.3. Electroporation
In this method DNA is inserted into the cells after which they are exposed to high
voltage electrical pulse which is believed to produce transient pores within the
plasma lemma. This approach requires the additional effort of weakening the cell
wall as it acts as an effective barrier against entry of DNA into cell cytoplasm hence,
it requires mild enzymatic treatment.
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3.2. M echanism of Action
Since almost all human pathogens invade at mucosal surfaces via urogenital,
respiratory and gastrointestinal tracts as their leading path of entry into the body.
Thus, foremost and prime line of the defense mechanism is mucosal immunity
(Moss, Cutts et al. 1999). The most efficient path of mucosal immunization is
oral route because oral vaccines are able to produce mucosal immunity, antibody
mediated immune response and cell mediated immune response. As an advantage
orally administered antigen containing plant vaccine do not get hydrolysed by
gastric enzymes due to tough outer wall of the plant cell. Transgenic plants
containing antigens act by the process of bioencapsulation i.e., outer rigid cell
wall and are finally hydrolysed and released in the intestines. The released
antigens are taken up by M cells in the intestinal lining that are placed on Payer’s
patches and gut-associated lymphoid tissue (GALT). These are further passed on
to macrophages and locallymphocyte populations, producing serum IgG, IgE
responses, local IgA response and memory cells, that rapidly counterbalance the
attack by the real infectious agent (Mercenier, Wiedermann et al. 2001)
Figure 1: Mechanism of edible vaccine.
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3.3. Advantages of Edible Vaccines
1. Edible vaccines have efficient mode of action for immunization, as they do not
require subsidiary elements to stimulate immune response.
2. Edible vaccine unlike traditional vaccines brings forth mucosal immunity.
3. Edible vaccines are comparatively cost effective, as they do not require cold
chain storage like traditional vaccines (Nochi, Takagi et al. 2007).
4. Edible vaccines offer greater storage opportunities as they seeds of transgenic
plants contain lesser moisture content and can be easily dried. In addition,
plants with oil or their aqueous extracts possess more storage opportunities.
5. Edible vaccines do not need sophisticated equipments and machines as they
could be easily grown on rich soils and the method is economical compared to
cell culture grown in fermenters.
6. Edible vaccines are widely accepted as they are orally administered unlike
traditional vaccines that are injectable. Thus, they eliminate the requirement of
trained medical personnel and the risk of contamination is reduced as they do
not need premises and manufacturing area to be sterilized (Streatfield, Jilka et
al. 2001).
7. Edible vaccines offer greater opportunity for second-generation vaccines by
integrating numerous antigens, which approach M cells simultaneously
8. Edible vaccines are safe as they do not contain heat-killed pathogens and hence
do not present any risk of proteins to reform into infectious organism.
9. Edible vaccine production process can be scaled up rapidly by breeding.
3.4. Limitations of Edible Vaccines
Following are some major drawbacks of edible vaccines,
Individual may develop immune tolerance to the particular vaccine protein or
peptide.
Dosage required varies from generation to generation and, plant to plant,
protein content, patient is age, weight, ripeness of the fruit and quantity of the
food eaten.
Edible vaccine administration requires methods for standardization of plant
material/product as low doses may produce lesser number of antibodies and
high doses are responsible immune tolerance.
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Edible vaccines are dependent on plant stability as certain foods cannot be
eaten raw (e.g. potato) and needs cooking that cause denaturation or weaken the
protein present in it (Moss, Cutts et al. 1999).
Edible vaccines are prone to get microbial infestation e.g. potatoes containing
vaccine can last long if stored at 4°C while a tomato cannot last long.
Proper demarcation line is necessary y between ‘vaccine fruit’ and ‘normal
fruit’ to avoid misadministration of vaccine, which can lead to vaccine
tolerance.
Edible vaccine function can be hampered due to vast differences in the
glycosylation pattern of plants and humans.
3.5. Applications
Cancertherapy
Several plants have been successfully engineered to generate monoclonal
antibodies that have been verified as effective cancer therapy agents. One
example is that of monoclonal body in case of soyabean (BR-96) is an efficient
agent that attacks doxorubicin responsible for breast cancer, ovarian cancer,
colon cancer and lung tumors (Prakash 1996).
Birth control
Administration of TMV produces protein that is found in Mousezona
pellucida (ZB3 protein) and is capable of preventing fertilization of eggs in
mice due to resulting antibodies (Prakash 1996).
Chloroplast transformation
As the chloroplast genome cannot be transmitted with in crops via usual cross
pollination due to its nature of maternal inheritance (Daniell, Khan et al.
2002). It may contribute to its transmission as well as accumulation in ample
quantities in the form of transgenic protein.
Role in autoimmune diseases
In concern with autoimmune diseases, scaling up of self- antigen production in
plants is underway in its developmental stage. Few of the diseases that are
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under study include; multiple sclerosis, rheumatoid arthritis, lupus and
transplant rejection. In one clinical study strain of mouse susceptible to diabetes
were fed with potatoes capable of expressing insulin and a protein called GAD
(glutamic acid decarboxylase), linked to CT-B subunit. It has been found out
that the protein proved successful in suppressing immune attack and delayed
the onset of high blood sugar level.
Recombinant drugs/proteins
Besides, being major producers of vaccines and antibodies, plant compositions
are altered by engineered viral inoculations to produce enzymes; drugs
(albumin, serum protease and interferon) e.g. glucocerebrosidase (hGC)
production in tobacco plants for treating Gaucher’s disease, Interleukin-10 to
treat Crohn’s disease This method of production is quite cheaper and reduces
the cost by thousand-fold (Prakash 1996). The process of recombinant
therapeutic protein production from plants has been commercialized as hirudin
which is an antithrombin-anti-viral protein that inhibits the HIV virus in vitro,
trichosanthin(ribosome in activator) and angiotensin-I (antihypersensitive
drug).
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4. FUTURE ASPECTS OF EDIBLES VACCINES
Resistance towards GM foods presents a threat to the rising future of edible
vaccines. Transgenic contamination is also a major concern which needs to be
addressed properly. Edible vaccines before launching in market for human
applications require certification by WHO in terms of its quality, efficiency and
environmental effect. Despite above concerns the future of edible vaccines is
reflected by enormous increase in land area used for cultivation of transgenic
crops from 1.7 to 44.2 million hectares from 1996 to 2000. Also the number of
countries growing them increased from 6 to 13 which predicted that transgenic
crops gained wide acceptance industrially as well as in developing countries.
Edible vaccines present good economics and technological benefits as more than
350 genetically engineered products are presently in progress in the United States
and Canada. In the near future edible vaccine against smallpox, anthrax, plague,
etc can be produced on a large scale (up to millions of doses) within a short span
of time.
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5. CONCLUSION
Edible plant-derived vaccines present a better possibility of safer and more
efficient immunization in the future. Limitations linked with traditional vaccines,
like production, distribution and delivery can be eliminated by the use of edible
vaccines through various immunization programs. Edible vaccines successfully
embraced the obstacles encountered in rising vaccine technology. Despite
restricted global access to health care and much attention still being paid towards
complex diseases like HIV, malaria, etc. The time is not so far when there is need
for an economical, safer and efficient delivery system to be developed at a larger
scale in the form of edible vaccines The ray of hope is based on assumption that
edible vaccines may be grown mostly in the developing countries which is
basically a fact as in reality they would be used in these countries. Hence, edible
vaccines provide a greater opportunity in the near future when no longer injectable
needles be used but a fruitful path may be available where an individual get
protected from diseases by simply eating a fruit.
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